A preliminary report on the exploration of salivary bacterial diversity by the multiplex SNaPshot assay.

Salivary bacterial community composition is associated with the host's internal and environmental factors, which have potential applications in forensic practice. The 16S rRNA gene sequencing is the most commonly used strategy for detecting salivary bacterial diversity; however, its platforms are not compatible with capillary electrophoresis (CE) platforms commonly used for forensic applications. Therefore, we attempted to detect the salivary bacterial diversity using a single nucleotide polymorphism (SNP) assay. Salivary bacterial diversity varies among diverse geographic locations, making it a potential supplementary biomarker for forensic geographic sourcing. To evaluate the performance of the multiplex SNaPshot assay, saliva samples from three geographic locations in China were analyzed using the multiplex SNaPshot assay and 16S rRNA gene sequencing. We screened SNPs from two high-relative-abundance salivary genera (Streptococcus and Veillonella) to construct a multiplex SNaPshot system that can be used on the CE platform. The stability and sensitivity of the multiplex SNaPshot system were also tested. A random forest classification model was used to classify samples from different regions to explore the ability of salivary bacteria to discriminate between geographic sources. Six bacterial SNPs were screened and a multiplex SNaPshot system was constructed. The stability results showed that the typing of salivary stains that were placed indoors for different days was not affected in this study. Two-thirds of mocked salivary stain samples showed more than 90% of typing results obtained for salivary stain samples with an input of 0.1 µl saliva. The results of principal coordinate analysis based on salivary bacterial diversity showed significant differences between samples from the three different geographic locations. The accuracy of the random forest classification was 66.67% based on the multiplex SNaPshot assay and 83.33% based on the 16S rRNA gene sequencing. In conclusion, this is the first attempt to detect salivary bacterial diversity using a multiplex SNaPshot bacterial SNP assay. The geographic difference in human salivary bacterial community composition was significant, as revealed by the multiplex SNaPshot assay; however, its performance in discriminating geographic sources was lower than that of 16S rRNA gene sequencing. This strategy based on bacterial SNP loci may favor the detection of human bacterial diversity in common forensic laboratories but requires further exploration in larger sample sizes and more bacterial SNP loci.

Social Work in Funeral Homes, a Unique Chinese Practice?

Social workers were introduced to funeral homes in China amid the transition and expansion of both the funeral home industry and the social work profession and are proving to play a valuable, though under-researched role in serving not just clients but also communities and funeral home staff. Funeral home social work fills gaps in after-death care and mental health and is distinct from palliative, hospice, end-of-life, and bereavement social work. Based on the experiences of funeral homes that employ social workers, this article argues that this innovation may bring new ideas to bridge some of the service gaps in after-death care in China and globally. This article outlines the support that will be needed from funeral homes, social work service agencies, and educational and research institutes to facilitate further development of funeral home mental health and social services and to promote the professionalization of funeral home social workers in China.

Partial loss of TDP-43 function causes phenotypes of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal neurological disease that causes motor neuron degeneration, progressive motor dysfunction, paralysis, and death. Although multiple causes have been identified for this disease, >95% of ALS cases show aggregation of transactive response DNA binding protein (TDP-43) accompanied by its nuclear depletion. Therefore, the TDP-43 pathology may be a converging point in the pathogenesis that originates from various initial triggers. The aggregation is thought to result from TDP-43 misfolding, which could generate cellular toxicity. However, the aggregation as well as the nuclear depletion could also lead to a partial loss of TDP-43 function or TDP-43 dysfunction. To investigate the impact of TDP-43 dysfunction, we generated a transgenic mouse model for a partial loss of TDP-43 function using transgenic RNAi. These mice show ubiquitous transgene expression and TDP-43 knockdown in both the periphery and the central nervous system (CNS). Strikingly, these mice develop progressive neurodegeneration prominently in cortical layer V and spinal ventral horn, motor dysfunction, paralysis, and death. Furthermore, examination of splicing patterns of TDP-43 target genes in human ALS revealed changes consistent with TDP-43 dysfunction. These results suggest that the CNS, particularly motor neurons, possess a heightened vulnerability to TDP-43 dysfunction. Additionally, because TDP-43 knockdown predominantly occur in astrocytes in the spinal cord of these mice, our results suggest that TDP-43 dysfunction in astrocytes is an important driver for motor neuron degeneration and clinical phenotypes of ALS.

Widespread aggregation of mutant VAPB associated with ALS does not cause motor neuron degeneration or modulate mutant SOD1 aggregation and toxicity in mice.

BACKGROUND: A proline-to-serine substitution at position-56 (P56S) of vesicle-associated membrane protein-associated protein B (VAPB) causes a form of dominantly inherited motor neuron disease (MND), including typical and atypical amyotrophic lateral sclerosis (ALS) and a mild late-onset spinal muscular atrophy (SMA). VAPB is an integral endoplasmic reticulum (ER) protein and has been implicated in various cellular processes, including ER stress, the unfolded protein response (UPR) and Ca2+ homeostasis. However, it is unclear how the P56S mutation leads to neurodegeneration and muscle atrophy in patients. The formation of abnormal VAPB-positive inclusions by mutant VAPB suggests a possible toxic gain of function as an underlying mechanism. Furthermore, the amount of VAPB protein is reported to be reduced in sporadic ALS patients and mutant SOD1G93A mice, leading to the hypothesis that wild type VAPB plays a role in the pathogenesis of ALS without VAPB mutations. RESULTS: To investigate the pathogenic mechanism in vivo, we generated human wild type (wtVAPB) and mutant VAPB (muVAPB) transgenic mice that expressed the transgenes broadly in the CNS. We observed robust VAPB-positive aggregates in the spinal cord of muVAPB transgenic mice. However, we failed to find an impairment of motor function and motor neuron degeneration. We also did not detect any change in the endogenous VAPB level or evidence for induction of the unfolded protein response (UPR) and coaggregation of VAPA with muVAPB. Furthermore, we crossed these VAPB transgenic mice with mice that express mutant SOD1G93A and develop motor neuron degeneration. Overexpression of neither wtVAPB nor muVAPB modulated the protein aggregation and disease progression in the SOD1G93A mice. CONCLUSION: Overexpression of VAPBP56S mutant to approximately two-fold of the endogenous VAPB in mouse spinal cord produced abundant VAPB aggregates but was not sufficient to cause motor dysfunction or motor neuron degeneration. Furthermore, overexpression of either muVAPB or wtVAPB does not modulate the course of ALS in SOD1G93A mice. These results suggest that changes in wild type VAPB do not play a significant role in ALS cases that are not caused by VAPB mutations. Furthermore, these results suggest that muVAPB aggregates are innocuous and do not cause motor neuron degeneration by a gain-of-toxicity, and therefore, a loss of function may be the underlying mechanism.

The C-terminal TDP-43 fragments have a high aggregation propensity and harm neurons by a dominant-negative mechanism.

TAR DNA binding protein 43 KD (TDP-43) is an essential gene that regulates gene transcription, mRNA splicing and stability. In amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two fatal neurodegenerative diseases, TDP-43 is fragmented, generating multiple fragments that include the C-terminal fragment of ∼25 KD. The role of these fragments in the pathogenesis of ALS and FTD is not clear. Here we investigated the aggregation propensity in various polypeptide regions of TDP-43 in mammalian cells and the effect of these fragments on cultured neurons. By expressing the full length and various TDP-43 fragments in motor neuron-derived NSC-34 cells and primary neurons, we found that both N- and C-terminal fragments of TDP-43 are prone to aggregate and the C-terminal end of RRM2 region is required, though not sufficient, for aggregation. The aggregation of the TDP-43 fragments can drive co-aggregation with the full-length TDP-43, consequently reducing the nuclear TDP-43. In addition, the TDP-43 fragments can impair neurite growth during neuronal differentiation. Importantly, overexpression of the full-length TDP-43 rescues the neurite growth phenotype whereas knockdown of the endogenous TDP-43 reproduces this phenotype. These results suggest that TDP-43 fragments, particularly the pathologically relevant C-terminal fragments, can impair neuronal differentiation by dominant-negatively interfering with the function of the full length TDP-43, thus playing a role in pathogenesis in ALS and FTD.

Infection with the insect virus Hz-2v alters mating behavior and pheromone production in female Helicoverpa zea moths.

The effect of Hz-2V virus infection on the reproductive physiology and behavior of infected Helicoverpa zea female moths was examined. In the absence of males, infected females exhibited calling behavior and called as often but for shorter periods on average than control females. As expected, control females mated with males for extend periods when they were present and did not call after mating, while virus-infected females made many frequent contacts with males and continued to call even after these contacts. Virus-infected females were found to produce five to seven times more pheromone than control females and attracted twice as many males as did control females in flight tunnel experiments. The ability of Hz-2V to alter the physiology and behavior of infected females observed here may serve to facilitate the transmission of virus in insect populations.

Horizontal transmission of Hz-2V by virus infected Helicoverpa zea moths.

Helicoverpa zea female moths productively infected with Hz-2V have malformed reproductive tissues and are sterile. Virus replication in infected females occurs primarily in the reproductive tissues and culminates with the accumulation of virus-filled vesicles, which form plugs of virus covering the reproductive openings of these insects. The location of this large concentration of virus particles at the terminal abdominal segment of infected females suggests that it may serve as a source of virus that can be transmitted horizontally between moths during mating. In mating experiments it was found that healthy males are attracted to and attempt to mate with infected females, and that these males are able transmit Hz-2V to healthy females during subsequent matings, giving rise to virus infected progeny.

Host plant effects on resistance to bifenthrin in silverleaf whitefly (Homoptera: Aleyrodidae).

Effects of host plants on resistance to bifenthrin in the silverleaf whitefly, Bemisia argentifolii Bellows & Perring, were determined by LC50 bioassay. In addition, inheritance of resistance to bifenthrin was investigated beginning with a single source of a bifenthrin-susceptible population. Overall, the resistance ratio between the bifenthrin-susceptible population and the selected bifenthrin-resistant population from the same source population was 915-fold after 1 yr in the greenhouse. Responses to bifenthrin among the susceptible and the resistant populations were changed when whiteflies were reared on three different host plants, i.e., cotton, cabbage, and squash. In the resistant populations, the LC50 value of whitefly fed on squash was increased as much as 7.5-fold, while the LC50 value of whitefly fed on cabbage was similar to cotton that served as the control plant. The host plant on which whiteflies feed appears to be an important factor in selection for resistance to bifenthrin, but these effects are crop specific. Based on an analysis using LC50 values of the reciprocal F1 cross on cotton, resistance of whitefly from a single-source whitefly population was inherited as an incompletely dominant factor. A model used to estimate loci numbers showed that resistance of whitefly to bifenthrin is probably controlled primarily by a few or a single locus. In addition, the difference in the ratio of LC50 values between males from unmated mother and males from mated mother was approximately fivefold, suggesting that insecticide resistance in whitefly males is in some way affected by mating.